Arylfluorosulfate compounds and methods

Abstract

A high-throughput screening methods for identifying candidate anticancer medicinal agents is described herein. The candidate anticancer medicinal agents are arylfluorosulfate compounds derived from phenolic compounds. The method involves in situ generation of the arylfluorosulfate compounds in multi-well plates by reaction of phenolic compounds in DMSO with a saturated solution of SO 2 F 2 dissolved in a solvent such as acetonitrile, in the presence of an organic base, followed by reaction of generated fluoride ion with trimethylsilanol to form volatile trimethylsilyl fluoride. Solvents, organic base, and silyl compounds are then removed, in vacuo, to afford the arylfluorosulfate compounds suitable for biological screening in cancer cell lines without further purification.

What is claimed is: 
     
       1. A high-throughput screening method for identifying anticancer medicinal agents comprising an arylfluorosulfate functional group, the method comprising the sequential steps of:
 (a) distributing solutions of phenolic compounds into wells of a first multi-well plate with one phenolic compound per well, wherein each phenolic compound comprises one or more OH substituent on an aromatic moiety; 
 (b) adding to each well of said first multi-well plate an organic base and a saturated solution of SO 2 F 2 , thereafter sealing each well, thereby initiating reactions between said OH substituents of the phenolic compounds with the SO 2 F 2  and forming candidate arylfluorosulfate anticancer agents therefrom; 
 (c) adding trimethylsilanol to each well in the first multi-well plate after the reaction in step (b) is complete to convert fluoride ion formed from the reaction of the phenolic compound and the SO 2 F 2  into trimethylsilyl fluoride; 
 (d) evaporating, in vacuo, solvents, the organic base, and trimethylsilyl compounds present in the wells to leave a residue of a candidate arylfluorosulfate anticancer agent in each well; 
 (e) dissolving each residue in dimethylsulfoxide to form solutions of the candidate arylfluorosulfate anticancer agents; 
 (f) adding selected concentrations of the candidate arylfluorosulfate anticancer agent solutions from step (e) to cancer cell cultures distributed in wells of a second multi-well plate; 
 (g) incubating the cell cultures in the second multi-well plate under conditions suitable for viability or growth of the cancer cells; and 
 (h) assessing the effect of the candidate arylfluorosulfate anticancer medicinal agents on the viability or growth of the cancer cells. 
 
     
     
       2. The method of  claim 1 , wherein the solutions in step (a) comprise the phenolic compounds dissolved in dimethylsulfoxide. 
     
     
       3. The method of  claim 1 , wherein the organic base in step (b) comprises triethylamine. 
     
     
       4. The method of  claim 1 , wherein the organic base in step (b) comprises diisopropylethylamine. 
     
     
       5. The method of  claim 1 , wherein the saturated solution of SO 2 F 2  in step (b) comprises SO 2 F 2  dissolved in acetonitrile. 
     
     
       6. The method of  claim 1 , wherein the reactions in step (b) are continued for about 12 hours before step (c). 
     
     
       7. The method of  claim 1 , wherein each solution of phenolic compound in step (a) comprises about 0.1 μmol of the phenolic compound dissolved in about 10 μL of DMSO. 
     
     
       8. The method of  claim 7 , wherein the saturated solution of SO 2 F 2  in step (b) comprises SO 2 F 2  dissolved in acetonitrile. 
     
     
       9. The method of  claim 8 , wherein about 100 μL of the saturated solution of SO 2 F 2  is added to each well in step (b). 
     
     
       10. The method of  claim 1  wherein the cancer cell cultures in step (f) comprise A549 adenocarcinomic human alveolar basal epithelial cells. 
     
     
       11. The method of  claim 1 , wherein the cancer cell cultures in step (f) comprise MCF-7 breast cancer cells. 
     
     
       12. The method of  claim 1 , wherein the cancer cell cultures in step (f) comprise SKBR3 breast cancer cells. 
     
     
       13. The method of  claim 1 , wherein the cancer cell cultures in step (f) comprise HT-29 colon cancer cells. 
     
     
       14. The method of  claim 1 , wherein at least two different concentrations of each candidate arylfluorosulfate anticancer agent are added to the cancer cell cultures in step (f). 
     
     
       15. The method of  claim 1 , wherein at least five different concentrations of each candidate arylfluorosulfate anticancer agent are added to the cancer cell cultures in step (f). 
     
     
       16. The method of  claim 1 , wherein the cancer cell cultures in step (f) are incubated at about 37° C. with 5% CO 2  for about 24 hours. 
     
     
       17. The method of  claim 1 , further comprising adding dimethylsulfoxide alone to some of the cell cultures in the second multi-well plate in step (f) as controls. 
     
     
       18. The method of  claim 17 , wherein the assessing in step (h) comprises calculating a growth inhibition percentage for each candidate arylfluorosulfate anticancer agent by comparing cell growth in the cultures containing the candidate arylfluorosulfate anticancer agent to cell growth of the controls. 
     
     
       19. The method of  claim 18 , wherein step (f) comprises adding at least five different concentrations of the candidate arylfluorosulfate anticancer agents to the cancer cell cultures; and the assessing in step (h) comprises constructing a dose-response curve from the cancer cell growth at each concentration of each candidate arylfluorosulfate anticancer agent. 
     
     
       20. A high-throughput screening method for identifying anticancer medicinal agents comprising an arylfluorosulfate functional group, the method comprising the sequential steps of:
 (a) distributing solutions comprising about 0.1 μmol of phenolic compounds dissolved in dimethylsulfoxide into wells of a first multi-well plate with one phenolic compound per well, wherein each phenolic compound comprises one or more OH substituent on an aromatic moiety and each solution comprises about 1 μmol of the phenolic compound; 
 (b) adding to each well of said first multi-well plate about 10 equivalents of an organic base and about 100 μL of a saturated solution of SO 2 F 2 , thereafter sealing each well, thereby initiating reactions between said OH substituents of the phenolic compounds with the SO 2 F 2  and forming candidate arylfluorosulfate anticancer agents therefrom; 
 (c) adding an excess of trimethylsilanol to each well in the first multi-well plate after the reaction in step (b) is complete to convert fluoride ion formed from the reaction of the phenolic compound and the SO 2 F 2  into trimethylsilyl fluoride; 
 (d) evaporating, in vacuo, dimethylsulfoxide, the organic base, and trimethylsilyl compounds present in the wells to leave a residue of a candidate arylfluorosulfate anticancer agent in each well; 
 (e) dissolving each residue in about 10 μL of dimethylsulfoxide to form solutions of the candidate arylfluorosulfate anticancer agents; 
 (f) adding at least two concentrations of the candidate arylfluorosulfate anticancer agent solutions from step (e) in the range of about 2×10 −7  μM to about 20 μM to cancer cell cultures distributed in wells of a second multi-well plate at cell concentrations of about 3000 to about 5000 cells per well; 
 (g) incubating the cell cultures in the second multi-well plate under conditions suitable for viability or growth of the cancer cells; and 
 (h) assessing the effect of the candidate arylfluorosulfate anticancer medicinal agents on the viability or growth of the cancer cells.